A Standardized DNA Variant Scoring System for Pathogenicity Assessments in Mendelian Disorders.

We developed a rules-based scoring system to classify DNA variants into five categories including pathogenic, likely pathogenic, variant of uncertain significance (VUS), likely benign, and benign. Over 16,500 pathogenicity assessments on 11,894 variants from 338 genes were analyzed for pathogenicity based on prediction tools, population frequency, co-occurrence, segregation, and functional studies collected from internal and external sources. Scores were calculated by trained scientists using a quantitative framework that assigned differential weighting to these five types of data. We performed descriptive and comparative statistics on the dataset and tested interobserver concordance among the trained scientists. Private variants defined as variants found within single families (n = 5,182), were either VUS (80.5%; n = 4,169) or likely pathogenic (19.5%; n = 1,013). The remaining variants (n = 6,712) were VUS (38.4%; n = 2,577) or likely benign/benign (34.7%; n = 2,327) or likely pathogenic/pathogenic (26.9%, n = 1,808). Exact agreement between the trained scientists on the final variant score was 98.5% [95% confidence interval (CI) (98.0, 98.9)] with an interobserver consistency of 97% [95% CI (91.5, 99.4)]. Variant scores were stable and showed increasing odds of being in agreement with new data when re-evaluated periodically. This carefully curated, standardized variant pathogenicity scoring system provides reliable pathogenicity scores for DNA variants encountered in a clinical laboratory setting.

Pulmonary Arterial Hypertension: A Current Perspective on Established and Emerging Molecular Genetic Defects.

Pulmonary arterial hypertension (PAH) is an often fatal disorder resulting from several causes including heterogeneous genetic defects. While mutations in the bone morphogenetic protein receptor type II (BMPR2) gene are the single most common causal factor for hereditary cases, pathogenic mutations have been observed in approximately 25% of idiopathic PAH patients without a prior family history of disease. Additional defects of the transforming growth factor beta pathway have been implicated in disease pathogenesis. Specifically, studies have confirmed activin A receptor type II-like 1 (ACVRL1), endoglin (ENG), and members of the SMAD family as contributing to PAH both with and without associated clinical phenotypes. Most recently, next-generation sequencing has identified novel, rare genetic variation implicated in the PAH disease spectrum. Of importance, several identified genetic factors converge on related pathways and provide significant insight into the development, maintenance, and pathogenetic transformation of the pulmonary vascular bed. Together, these analyses represent the largest comprehensive compilation of BMPR2 and associated genetic risk factors for PAH, comprising known and novel variation. Additionally, with the inclusion of an allelic series of locus-specific variation in BMPR2, these data provide a key resource in data interpretation and development of contemporary therapeutic and diagnostic tools.

Improved detection of cystic fibrosis mutations in infertility patients with DNA sequence analysis.

BACKGROUND: Accurate determination of mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene is critical for genetic counselling and treatment of obstructive azoospermia. Of concern is that detection rates with routine CFTR mutation panels vary widely depending on patient ancestry; and such panels have limited value for azoospermic patients, who are more likely to carry rare mutations. An alternative approach offers comprehensive, CFTR mutation analysis by a DNA sequence method. We investigated whether this method could improve CFTR detection rates in men with obstructive azoospermia in a prospective study of men with obstructive azoospermia and their partners who were referred for genetic counselling and testing at one of two institutions. METHODS: Sixteen patients with congenital absence of the vas deferens (CAVD, n = 14) or idiopathic obstructive azoospermia (n = 2) were studied. DNA from all patients was analysed for mutations by the DNA sequence method. In addition to this method, six men underwent CFTR analysis by a common 25 or 31 mutation panel coupled with poly T analysis. In 10 subjects, common mutation panel findings were inferred from DNA sequence method results. RESULTS: Overall, 12/16 (75%) azoospermic patients had one or more CFTR mutations and/or 5T alleles, including 12 mutations in 10 patients (two compound heterozygotes) and seven 5T alleles in six patients (one homozygote). The sequence method detected all mutations and three variants of unknown significance. By comparison, the common mutation panels detected only 3/12 mutations (25%) and 0/3 variants. CONCLUSION: The DNA sequence method detects more CFTR mutations than common mutation panels. Given the serious, clinical consequences of transmitting such mutations, this study underscores the importance of accurate, CFTR mutation detection in men with obstructive azoospermia and their partners.